Granger-Schnarr M, Daune M P, Fuchs R P
J Mol Biol. 1986 Aug 5;190(3):499-507. doi: 10.1016/0022-2836(86)90018-5.
The mismatch repair system of Escherichia coli is known to contribute to the fidelity of the replicational process. This system involves the functions of mutH, mutL, mutS and mutU (uvrD) loci which recognize mispaired bases as a consequence of errors due to the polymerase itself. Chemical modifications of DNA have also been suspected to create mispaired bases which, if the mispaired bases are removed, will lead to mutations by frameshift. Using the pBR322 plasmid DNA modified by the ultimate carcinogen N-acetoxy-N-2-acetylaminofluorene (N-Aco-AAF) we have investigated this possibility in a forward mutational assay (tetracycline sensitivity). This fluorene derivative has been shown to induce predominantly frameshift mutations. Our results show that: The sensitivity of the deficient strains mutH, mutL and mutS to the AAF adducts is similar to that of the corresponding wild-type strain. However, the mutU strain appears much more sensitive to those adducts although less than a uvrA, B or C-deficient strain. This suggests that the mutU gene product is involved in the repair of AAF adducts. For the four mut deficient strains, and as it was shown with the wild-type strain, AAF adducts induced mutations to tetracycline sensitivity are only observed when the SOS system of the host bacteria is induced by irradiation of the cells prior to transformation with the modified plasmid. The mutation frequencies depend upon the ultraviolet light doses and similar maxima were found for the four mut strains and the corresponding wild-type strain. In agreement with the results obtained with wild-type or uvrA strains we observe that AAF adducts induce mostly frameshift mutations in the mut strains. Two types of hot spots of mutagenesis were described in wild-type and uvrA strains occurring either at repetitive sequences or at sequences of the type 5' G-G-C-G-C-C 3' (NarI restriction enzyme recognition sequence). While the second type of mutational hot spot does exist in the mismatch repair-deficient strains, we observe that the repetitive sequences are no longer hot spots of mutations in these strains, suggesting that the mismatch repair protein complex is involved in the establishment of AAF-induced frameshift mutations at repetitive sequences.
已知大肠杆菌的错配修复系统有助于提高复制过程的保真度。该系统涉及mutH、mutL、mutS和mutU（uvrD）基因座的功能，这些基因座可识别由于聚合酶本身的错误而导致的错配碱基。DNA的化学修饰也被怀疑会产生错配碱基，如果去除这些错配碱基，将通过移码导致突变。我们使用由终极致癌物N-乙酰氧基-N-2-乙酰氨基芴（N-Aco-AAF）修饰的pBR322质粒DNA，在正向突变试验（四环素敏感性）中研究了这种可能性。这种芴衍生物已被证明主要诱导移码突变。我们的结果表明：mutH、mutL和mutS缺陷菌株对AAF加合物的敏感性与相应的野生型菌株相似。然而，mutU菌株对这些加合物似乎更敏感，尽管不如uvrA、B或C缺陷菌株敏感。这表明mutU基因产物参与了AAF加合物的修复。对于这四种mut缺陷菌株，正如在野生型菌株中所显示的那样，只有当宿主细菌的SOS系统在使用修饰质粒转化之前通过细胞照射诱导时，才会观察到AAF加合物诱导的对四环素敏感性的突变。突变频率取决于紫外线剂量，并且在四种mut菌株和相应的野生型菌株中发现了相似的最大值。与野生型或uvrA菌株获得的结果一致，我们观察到AAF加合物在mut菌株中主要诱导移码突变。在野生型和uvrA菌株中描述了两种诱变热点，分别出现在重复序列或5' G-G-C-G-C-C 3'类型的序列（NarI限制酶识别序列）处。虽然第二种类型的突变热点确实存在于错配修复缺陷菌株中，但我们观察到重复序列在这些菌株中不再是突变热点，这表明错配修复蛋白复合物参与了在重复序列处建立AAF诱导的移码突变。
